Differential toxicity of ganciclovir for rat neurons and astrocytes in primary culture following adenovirus-mediated transfer of the HSVtk gene.

The toxicity of the suicide HSVtk gene approach is known to be targeted to DNA synthesis and, consequently, to dividing cells. This system is therefore useful for the treatment of brain tumors which contain dividing cells surrounded by a quiescent normal tissue. Adenoviruses are efficient vectors for the transfer of the HSVtk gene into the tumor but this can lead to the transduction of quiescent cells. In this study, we focused on the toxicity of the HSVtk/ganciclovir treatment for the two main cell types of the normal brain: astrocytes and neurons. Astrocytes and neurons in primary culture were infected by an adenoviral vector bearing the HSVtk gene (Ad.tk) and cells were exposed to different concentrations of ganclclovir. After 5 days of treatment, an MTT test measured a dramatic decrease in cell viability for treated astrocytes while a small decrease in cell viability was observed for neurons treated in the same experimental conditions. The differential toxicity of the HSVtk/ganciclovir treatment was also observed in cocultures of astrocytes and neurons: an immunocytochemical analysis of the treated cells showed major morphological modifications for astrocytes but not for neurons. Furthermore, our data suggest that a bystander effect is able to kill all the astrocytes while neurons from the same culture remain unaffected.

Role of virus receptor-bearing endothelial cells of the blood-brain barrier in preventing the spread of mouse hepatitis virus-A59 into the central nervous system.

BALB/c mice develop a neurologic demyelinating disease after inoculation of mouse hepatitis virus (MHV), strain A59, by the intracranial, but not by the intraperitoneal route. To determine the mechanisms that prevent virus spreading through the blood-brain barrier, we analyzed expression of MHVR, a glycoprotein that serves as receptor for mouse hepatitis virus on endothelial cells of cerebral blood vessels. Our results indicated that MHVR was strongly expressed on the endoluminal pole of these cells. In addition, a direct virus binding assay showed that mouse hepatitis virus was able to bind endothelial cells via this receptor. Despite this expression of a functional viral receptor, in normal mice infected with mouse hepatitis virus by the contra-peritoneal route, no in vivo viral replication could be detected in endothelial cells from the brain, contrasting with the equivalent cells from the liver. However, shortly after i.v. administration of sodium dodecylsulfate detergent to the mice, virus infection of some cerebral endothelial cells was detected in a few mice. As a consequence of detergent treatment, virus infection was able to cross the blood-brain barrier. These results suggest that the protective role of the blood-brain barrier against spreading of mouse hepatitis virus A59 into the central nervous system is determined by a specific restriction of viral entry into the endothelial cells of cerebral origin.